HTRF术语详解 详细 >>
FRET：指荧光共振能量转移，在供体和受体相互靠得很近时，光子能从一个受激发的荧光团（供体）转移到另一个荧光团（受体），并使后者发出荧光。
HTRF：均相时间分辨荧光，Homogeneous Time-Resolved Fluorescence ）是用来检测纯液相体系中待测物的一种常用方法。
背景（Background）：指在没有加入compound时检测到的信号值。
荧光受体（acceptor）：指在荧光团受到激光激发时，能发出特定波长的发射光的基团。HTRF®的能量受体均可为XL665和d2。
资料下载 更多 >>
- New HTRF cellular platform for cell surface receptors' study and screening
- Homogeneous Time-Resolved Fluorescence Part1:Methodological aspects
- HTRF GPCR Solutions:Gi,Gs,Gq receptor screening using a single technology
- HTRF Compatible Readers:Unleash your lab's capacity to read HTRF technology
- Forget Elisa use HTRF biomarkers
Time-resolved Fluorescence Resonance Energy Transfer (TR-FRET) to Analyze the Disruption of EGFR/HER2 Dimers: A new method to evaluate the efficiency of targeted therapy using monoclonal antibodies.
By: Gaborit N, Larbouret C, Vallaghe J, Peyrusson F, Bascoul-Mollevi C, Crapez E, Azria D, Chardès T, Poul MA, Mathis G, Bazin H, Pèlegrin A.
IRCM, Institut de Recherche en Cancérologie de Montpellier, INSERM, U896, Université Montpellier, CRLC Val d'Aurelle Paul Lamarque (Montpellier, France).
noncology, simultaneous inhibition of epidermal growth factor receptor (EGFR) and HER2 by monoclonal antibodies (mAbs) is an efficient therapeutic strategy but the underlying mechanisms are not fully understood. Here, we describe a time-resolved fluorescence resonance energy transfer (TR-FRET) method to quantify EGFR/HER2 heterodimers on cell surface to shed some light on the mechanism of such therapies. First, we tested this antibody-based TR-FRET assay in NIH/3T3 cell lines that express EGFR and/or HER2 and in various tumor cell lines. Then, we used the antibody-based TR-FRET assay to evaluate in vitro the effect of different targeted therapies on EGFR/HER2 heterodimers in the ovarian carcinoma cell line SKOV-3. A simultaneous incubation with Cetuximab (anti-EGFR) and Trastuzumab (anti-HER2) disturbed EGFR/HER2 heterodimers resulting in a 72% reduction. Cetuximab, Trastuzumab or Pertuzumab (anti-HER2) alone induced a 48, 44, or 24% reduction, respectively. In contrast, the tyrosine kinase inhibitors Erlotinib and Lapatinib had very little effect on EGFR/HER2 dimers concentration. In vivo, the combination of Cetuximab and Trastuzumab showed a better therapeutic effect (median survival and percentage of tumor-free mice) than the single mAbs. These results suggest a correlation between the extent of the mAb-induced EGFR/HER2 heterodimer reduction and the efficacy of such mAbs in targeted therapies. In conclusion, quantifying EGFR/HER2 heterodimers using our antibody-based TR-FRET assay may represent a useful method to predict the efficacy and explain the mechanisms of action of therapeutic mAbs, in addition to other commonly used techniques that focus on antibody-dependent cellular cytotoxicity, phosphorylation, and cell proliferation.
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